The long term goal of this project is to clone, sequence, express and map the distribution of nutrient transporters in the blood-brain and blood- cerebrospinal-fluid barriers. The endothelial cells of the brain capillaries regulate the uptake of nutrients into the brain, and the epithelial cells of the choroid plexus clear the cerebrospinal fluid of many metabolites and drugs. Relatively little is known about the transporters involved in these vital processes. We will clone blood- brain barrier transporters using two different strategies. The first is to determine if the transporters already cloned from other tissues are expressed at the blood-brain-barrier. A candidate clone for brain capillaries is the renal Na+/myo-mositol cotransporter, since isolated brain microvessels transport this important phospholipid precursor, and a candidate for the choroid plexus is the renal Na+/nucleoside cotransporter. Northern and Western blots, in situ hybridization, and immunocytochemical techniques will be used to map the distribution of candidate clones at the blood-brain interface. The second strategy is to use expression cloning techniques to isolate specific brain capillary and choroid plexus transporters, e.g. The L-type amino acid and Na+- dependent proline transporters. The objective is to use the tools of cell and molecular biology to obtain a detailed understanding of nutrient and metabolite transport between the blood and brain.